Mapping of interleukin-5 functional domains and creation of a functional interleukin-5 monomer demonstrate that all structural features necessary for interleukin-5 biological activity are contained within a single helical bundle motif
Abstract
The cytokine interleukin-5 (IL-5) specifically induces the maturation and activation of eosinophils, which are important in normal host defense and allergic disease. Crucial to the development of specific modulators of IL-5 is a comprehensive understanding of the interaction of IL-5 with its receptor complex, which consists of alpha and beta subunits. IL-5 is a unique member of the short chain subfamily of helical bundle cytokines. IL-5 is a glycosylated, disulfide-linked, interdigitating homodimer, which contains two canonical helical bundle motifs. This distinguishes IL-5 from the other subfamily members, which fold unimolecularly into a single helical bundle. I undertook a systematic approach to identify functional domains on IL-5 and solve the enigma surrounding its unique dimeric configuration. A library of neutralizing monoclonal antibodies (mAb) was generated and characterized. Two noncompetitive anti-IL-5 mAb were used to develop a highly sensitive sandwich ELISA capable of quantitating IL-5 in biological fluids. The anti-IL-5 mAb were also used to map functional domains on IL-5. These results predicted regions which engaged the IL-5 receptor. Each functional domain was defined twice on the IL-5 dimer. However, the functional significance of each domain pair could not be established. To determine whether a single helical bundle of IL-5 was independently functional or if IL-5 required both helical bundles for bioactivity, I created a novel monomer, which folded unimolecularly into a single IL-5 helical bundle motif. Generation of this cytokine, designated mono5, relied on the hypothesis that the shortened loop 3 of IL-5 physically restricts unimolecular completion of a single helical bundle. Structural and functional analyses demonstrated that mono5 was indeed a monomer that possessed full biological activity comparable to native IL-5 at saturating concentrations. Although slight tertiary structural differences were identified, the full functional capacity of mono5 demonstrated for the first time that, all structural features necessary for IL-5 biological activity are contained within a single helical bundle motif. The creation of biologically active mono5 revolutionizes the understanding of IL-5 functional structure. Furthermore, my loop 3 hypothesis is applicable to the entire family of helical bundle cytokines and may lead to the creation of other novel proteins.
Subject Area
Immunology
Recommended Citation
Dickason, Richard Ray, "Mapping of interleukin-5 functional domains and creation of a functional interleukin-5 monomer demonstrate that all structural features necessary for interleukin-5 biological activity are contained within a single helical bundle motif" (1996). Texas Medical Center Dissertations (via ProQuest). AAI9619583.
https://digitalcommons.library.tmc.edu/dissertations/AAI9619583